Castration-resistant prostate cancer (CRPC) kills ~28,000 American men each year. The standard-of-care clinical therapies for advanced prostate cancer (PCa) have been androgen deprivation therapy (ADT) and/or chemotherapies (CT). ADT, aiming to block androgen synthesis (e.g., abiraterone) or to inhibit androgen receptor (AR) signaling (e.g., enzalutamide), achieves impressive short-term clinical effects by de-bulking primary tumor and reducing serum PSA. The reported survival benefit, however, is measured ONLY in months and most `castrated' patients eventually develop CRPC. Similarly, CT drugs including taxanes (docetaxel and cabazitaxel) and cisplatin have been used to treat advanced and recurrent PCa but resistance rapidly develops. The cellular origin and molecular mechanisms underpinning the emergence, sustenance, and progression of CRPC and CT-resistant PCa remain poorly understood. Systematic studies from our lab over the past 16 years on dissecting PCa cell heterogeneity and plasticity have pinpointed a population of phenotypically undifferentiated PCa stem cells (PCSCs) that frequently lack AR and are largely dormant, which, together, render them INHERENTLY resistant to both ADT and CT. Importantly, PCSCs frequently become the predominant cell population in therapy-resistant tumors and may function as a cell-of-origin as well as tumor-propagating cells for CRPC. These observations lead to our overarching hypothesis that combinatorial targeting of bulk PCa cells with ADT or CT, and PCSCs with novel strategies, will inhibit both cancer cell stemness and cellular plasticity, and achieve superior therapeutic efficacies and prevent/overcome therapy resistance. We test this hypothesis with 3 Specific Aims. Aim 1): To test the hypothesis that combining enzalutamide with Venetoclax (a newly and the ONLY FDA- approved BCL-2 inhibitor) will delay/prevent CRPC. Our recent xenograft modeling coupled with RNA-Seq and experimental therapies has pinpointed BCL-2 as a critical PCSC survival factor and a driver of CRPC. Aim 2): To test the hypothesis that combining enzalutamide with anti-PCSC microRNAs will delay/prevent CRPC. We have demonstrated that several tumor-suppressive miRNAs, including miR-34a and miR-141 are critical negative regulators of PCSCs and potently inhibit PCa metastasis and extend animal survival. Aim 3): To test the hypothesis that combining CT with anti-PCSC microRNAs or anti-BCL-2 will significantly extend the therapeutic efficacy of chemotherapies. These aims will be accomplished by combining extensive xenograft/PDX modeling and therapies with in-depth mechanistic studies. Accomplishment of the goals should help establish important principles, validate novel hypotheses, elucidate the molecular underpinnings of PCa subpopulation dynamics in response to treatments, and identify novel therapeutics and therapeutic regimens against different PCa subpopulations. These should facilitate rapid translation of our preclinical knowledge to clinical trials.